Inquiry in Action

Improving Tools for Gene Repair

Innovations in enzyme development made by the Scharenberg and Rawlings labs and their partners at the University of Washington and Fred Hutchinson Cancer Research Center move researchers closer to making precision gene manipulation a therapeutic reality.

Genetic disease is a major treatment challenge in medicine. Problems that result from genetic differences – whether hereditary or acquired – remain constant throughout an individual’s life span, often diminishing one’s quality of life and requiring ongoing treatment.

That is unless … the problematic gene could be modified to correct the root cause of the trouble. Developing the knowledge and technology to do this is one of the big pushes in biomedicine today.

“No other center in the country has the expertise to build the type of enzymes Andy’s team has. They were first structurally identified here. And the vast majority of the engineering has happened here.”~ Dr. Dave Rawlings When the field of gene therapy began about 30 years ago, the paradigm was to insert a replacement gene somewhere at random within the genome and leave the defective gene in place. Though successful for some applications, this approach has some significant drawbacks, from causing genes to behave erratically to damaging neighboring genes. So researchers began to wonder: how can we fix a gene?

A new approach to gene therapy

Drs. Andy Scharenberg and Dave Rawlings are pioneers in the emerging field of gene repair. This more precise and flexible approach to gene therapy has applications for healthcare as well as agricultural biotechnology. As co-directors of the Northwest Genome Engineering Consortium, Scharenberg and Rawlings lead a multi-disciplinary team that includes seven laboratories at the Seattle Children’s Research Institute, the University of Washington, and the Fred Hutchinson Cancer Research Center. They work toward a shared vision: developing the genetic tools needed to modify genes in a controlled and precise way to change or disrupt their function without affecting the rest of the genome.

The consortium has two primary thrusts: • Developing the enzymes, known as homing endonucleases, that bind to and cut a sequence of DNA precisely where the modification needs to be made.• Developing the means of delivering these enzymes to the cells.

“It’s taken us five years to build the tools we need to break the DNA where we want to,” says Scharenberg, who led a four-laboratory team to develop the homing endonucleases.

More precise tools

The enzymes created by the homing endonuclease development team are better suited for therapeutic purposes than those being developed elsewhere because:• They are smaller, making them easier to deliver to the gene that needs to be modified. • They are able to recognize DNA more specifically, enabling them to find their intended target along the genome.• They are not repetitive, making it easier for the enzyme to work with both the DNA it needs to modify and the viral technology being developed to deliver it.

They are also more complicated and painstaking to build: The team developed the enzymes by gathering lots of information about how alterations in naturally occurring homing endonucleases influence the kinds of DNA they target, bind to, and cut. Using that information – and some very sophisticated expertise and technology from the fields of computational protein design, biophysics, cell biology, bioinformatics and immunology – the team was able to create enzymes that can break the DNA precisely where needed.

“No other center in the country has the expertise to build the type of enzymes Andy’s team has,” says Rawlings. “They were discovered in Seattle. They were first structurally identified here. And the vast majority of the engineering of these enzymes has happened here.”

Moving from basic science discovery to real-world viability

Translating these innovative technologies from lab to patients is the next step for the teams. Their initial focus is on disrupting gene function to provide therapeutic benefit. They are pursuing promising applications that include treating blood disorders like sickle cell disease and other hemoglobin disorders, turning off genes in T cells to improve their ability to kill tumor cells, and disrupting the CCR5 receptor on cells that enables the HIV virus to enter them.

The teams will also continue working to develop the additional knowledge and tools needed to seamlessly fix the DNA of defective genes. “The creativity and fun of scientific discovery is always tempered by the fact that I’m a doctor,” says Scharenberg. “I’ve been working for the better part of my life to get something to patients that improves their lives. Everyone dreams about being part of a team that does this. It’s taken seven teams many years to make these tools. Now it’s time to take them and really make something.”

"The creativity and fun of scientific discovery is always tempered by the fact I'm a doctor. I've been working for the better part of my life to get something to patients that improves their lives. It's taken seven teams many years to make these tools. Now it's time to take them and really make something." ~Dr. Andy Scharenberg

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